Differences in intracellular DNA ligation after microinjection and transfection.
Abstract
Differences in intracellular DNA ligation after microinjection and transfection. J J Kopchick and D W Stacey ABSTRACT An uninterrupted avian sarcoma viral genome terminated by viral long terminal repeat sequences was cloned into a pBR322 plasmid. After introduction into a cultured avian cell, transcription of either the circular plasmid molecule or one linearized within the pBR322 sequences could initiate and terminate at long terminal repeat sequences, yielding full-sized viral RNA. A plasmid DNA molecule linearized by cleavage within the viral pol gene, on the other hand, would have to undergo ligation to yield full-sized viral RNA. Microinjection of each of these three types of DNA into the nuclei of quail cells promoted the release of similar virus titers, indicating that the plasmid DNA cleaved within the viral pol gene had been efficiently and accurately ligated. When plasmid DNA was transfected into quail cells, circular and pBR322-cleaved molecules directed the synthesis of similar virus titers, indicating that they were similarly taken up and utilized by the cells. Compared with these results, plasmid DNA cleaved within the pol gene was reduced in activity over 95% after transfection. This reduction did not result from inefficient ligation but from the generation of mutations (of limited size) during ligation of the transfected molecules. Mutations were not observed after microinjection even into the cytoplasm. Consistent with these findings, transfected DNA termini were found to be joined regardless of their structure, whereas ligation after microinjection required that single-stranded protruding DNA termini be complementary. CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? « Previous | Next Article » Table of Contents This Article doi: 10.1128/MCB.4.2.240 Mol. Cell. Biol. February 1984 vol. 4 no. 2 240-246 » Abstract PDF Classifications Research Article Services Email this article to a colleague Similar articles in ASM journals Alert me when this article is cited Alert me if a correction is posted Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of MCB Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Load citing article information Citing articles via Web of Science Citing articles via Google Scholar Google Scholar Articles by Kopchick, J. J. Articles by Stacey, D. W. Search for related content PubMed PubMed citation Articles by Kopchick, J. J. Articles by Stacey, D. W. Related Content Load related web page information Social Bookmarking CiteULike Connotea Delicious Digg Facebook Google+ Mendeley Reddit StumbleUpon Twitter What's this? current issue January 2012, volume 32, issue 1 Spotlights in the Current Issue Architecture of the Yeast RNA Polymerase II Open Complex State and Regulation by TFIIF GATA-1 Establishes Cell-Type-Specific Autophagy as a Developmental Program Prickle Phosphorylation Regulates Its Localization and β-Catenin-Independent Wnt Signaling Alert me to new issues of MCB About MCB Subscribers Authors Reviewers Advertisers Inquiries from the Press Permissions & Commercial Reprints ASM Journals Public Access Policy MCB RSS Feeds 1752 N Street N.W. • Washington DC 20036 202.737.3600 • 202.942.9355 fax • journals@asmusa.org Print ISSN: 0270-7306 Online ISSN: 1098-5549 Copyright © 2011 by the American Society for Microbiology. For an alternate route to MCB .asm.org, visit: http://intl- MCB .asm.org | More Info» var gaJsHost = (("https:" == document.location.protocol) ? "https://ssl." : "http://www."); document.write(unescape("%3Cscript src='" + gaJsHost + "google-analytics.com/ga.js' type='text/javascript'%3E%3C/script%3E")); var pageTracker = _gat._getTracker("UA-5821458-11"); pageTracker._trackPageview();